PROJECT SUMMARY/ABSTRACT Epilepsy affects more than 3 million people in the United States. In addition to unprovoked seizures, cognitive decline and memory impairment are common comorbidities associated with epilepsy, but our knowledge in this area is limited. Our preliminary work suggests that an epilepsy associated ubiquitin E3 ligase Nedd4-2 mediates actin polymerization through promoting phosphorylation of an actin binding protein cofilin during the induction of long-term synaptic potentiation (LTP). LTP describes long?lasting increments of synaptic efficiency and is crucial for cognition and memory formation. In an effort to identify ubiquitination substrates of Nedd4-2 responsible for cofilin phosphorylation, we identified PAK3, an IDG-eligible understudied kinase, as a potential substrate of Nedd4-2 that contributes to cofilin phosphorylation during LTP. This pilot project is formulated to test our hypothesis that induction of LTP de-represses Nedd4-2 to stabilize PAK3 and subsequently induce cofilin phosphorylation and actin polymerization. In Aim 1, we will determine how PAK3 is ubiquitinated by Nedd4-2 and how this ubiquitination is disrupted by epilepsy associated mutations in Nedd4-2. We will also determine whether induction of LTP de-represses Nedd4-2 to allow PAK3 stabilization during LTP. In Aim 2, we will employ a recently developed optogenetic approach to rapidly stabilize PAK3 in Nedd4-2 conditional knockout (cKO) neurons during the induction of LTP, with the intention to restore cofilin phosphorylation and actin polymerization, and ultimately to restore LTP. We expect our project to further our knowledge of an IDG-eligible protein PAK3 in synaptic plasticity and help explain cognitive decline in epilepsy.